ATP1A3 dysfunction causes motor hyperexcitability and afterhyperpolarization loss in a dystonia model.

Mutations in the gene encoding the alpha3 Na+/K+-ATPase isoform (ATP1A3) lead to movement disorders that manifest with dystonia, a common neurological symptom with many different origins, but for which the underlying molecular mechanisms remain poorly understood. We have generated an ATP1A3 mutant mouse that displays motor impairments and a hyperexcitable motor phenotype compatible with dystonia. We show that neurons harboring this mutation are compromised in their ability to extrude raised levels of intracellular sodium, highlighting a profound deficit in neuronal sodium homeostasis.

STED super-resolution microscopy of mitochondrial translocases.

The mitochondrial translocases of the outer membrane (TOM) and of the inner membrane (TIM) act together to facilitate the import of nuclear-encoded proteins across the mitochondrial membranes. Stimulated Emission Depletion (STED) super-resolution microscopy enables the in situ imaging of such complexes in single cells at sub-diffraction resolution. STED microscopy requires only conventional sample preparation techniques and provides super-resolved raw data without the need for further image processing.

Endogenous BAX and BAK form mosaic rings of variable size and composition on apoptotic mitochondria.

One hallmark of apoptosis is the oligomerization of BAX and BAK to form a pore in the mitochondrial outer membrane, which mediates the release of pro-apoptotic intermembrane space proteins into the cytosol. Cells overexpressing BAX or BAK fusion proteins are a powerful model system to study the dynamics and localization of these proteins in cells. However, it is unclear whether overexpressed BAX and BAK form the same ultrastructural assemblies following the same spatiotemporal hierarchy as endogenously expressed proteins.

DNA-PAINT MINFLUX nanoscopy.

MINimal fluorescence photon FLUXes (MINFLUX) nanoscopy, providing photon-efficient fluorophore localizations, has brought about three-dimensional resolution at nanometer scales. However, by using an intrinsic on-off switching process for single fluorophore separation, initial MINFLUX implementations have been limited to two color channels. Here we show that MINFLUX can be effectively combined with sequentially multiplexed DNA-based labeling (DNA-PAINT), expanding MINFLUX nanoscopy to multiple molecular targets.

A missense mutation converts the Na,K-ATPase into an ion channel and causes therapy-resistant epilepsy.

The ion pump Na,K-ATPase is a critical determinant of neuronal excitability; however, its role in the etiology of diseases of the central nervous system (CNS) is largely unknown. We describe here the molecular phenotype of a Trp931Arg mutation of the Na,K-ATPase catalytic α1 subunit in an infant diagnosed with therapy-resistant lethal epilepsy. In addition to the pathological CNS phenotype, we also detected renal wasting of Mg.

Mitochondrial Protein Abundance Gradients Require the Distribution of Separated Mitochondria.

Mitochondria are highly dynamic organelles that interchange their contents mediated by fission and fusion. However, it has previously been shown that the mitochondria of cultured human epithelial cells exhibit a gradient in the relative abundance of several proteins, with the perinuclear mitochondria generally exhibiting a higher protein abundance than the peripheral mitochondria. The molecular mechanisms that are required for the establishment and the maintenance of such inner-cellular mitochondrial protein abundance gradients are unknown.

A MICOS-TIM22 Association Promotes Carrier Import into Human Mitochondria.

Mitochondrial membrane proteins with internal targeting signals are inserted into the inner membrane by the carrier translocase (TIM22 complex). For this, precursors have to be initially directed from the TOM complex in the outer mitochondrial membrane across the intermembrane space toward the TIM22 complex. How these two translocation processes are topologically coordinated is still unresolved.

Mic60 exhibits a coordinated clustered distribution along and across yeast and mammalian mitochondria.

Mitochondria are tubular double-membrane organelles essential for eukaryotic life. They form extended networks and exhibit an intricate inner membrane architecture. The MICOS (mitochondrial contact site and cristae organizing system) complex, crucial for proper architecture of the mitochondrial inner membrane, is localized primarily at crista junctions.

ROMO1 is a constituent of the human presequence translocase required for YME1L protease import.

The mitochondrial presequence translocation machinery (TIM23 complex) is conserved between the yeast and humans; however, functional characterization has been mainly performed in yeast. Here, we define the constituents of the human TIM23 complex using mass spectrometry and identified ROMO1 as a new translocase constituent with an exceptionally short half-life. Analyses of a ROMO1 knockout cell line revealed aberrant inner membrane structure and altered processing of the GTPase OPA1.

Neuronal Aβ42 is enriched in small vesicles at the presynaptic side of synapses.

The amyloid β-peptide (Aβ) is a physiological ubiquitously expressed peptide suggested to be involved in synaptic function, long-term potentiation, and memory function. The 42 amino acid-long variant (Aβ42) forms neurotoxic oligomers and amyloid plaques and plays a key role in the loss of synapses and other pathogenic events of Alzheimer disease. Still, the exact localization of Aβ42 in neurons and at synapses has not been reported.

Experimental validation of predicted cancer genes using FRET.

Huge amounts of data are generated in genome wide experiments, designed to investigate diseases with complex genetic causes. Follow up of all potential leads produced by such experiments is currently cost prohibitive and time consuming. Gene prioritization tools alleviate these constraints by directing further experimental efforts towards the most promising candidate targets.

The MICOS component Mic60 displays a conserved membrane-bending activity that is necessary for normal cristae morphology.

The inner membrane (IM) of mitochondria displays an intricate, highly folded architecture and can be divided into two domains: the inner boundary membrane adjacent to the outer membrane and invaginations toward the matrix, called cristae. Both domains are connected by narrow, tubular membrane segments called cristae junctions (CJs). The formation and maintenance of CJs is of vital importance for the organization of the mitochondrial IM and for mitochondrial and cellular physiology.

TIM29 is a subunit of the human carrier translocase required for protein transport.

Hydrophobic inner mitochondrial membrane proteins with internal targeting signals, such as the metabolite carriers, use the carrier translocase (TIM22 complex) for transport into the inner membrane. Defects in this transport pathway have been associated with neurodegenerative disorders. While the TIM22 complex is well studied in baker's yeast, very little is known about the mammalian TIM22 complex.

Bax assembles into large ring-like structures remodeling the mitochondrial outer membrane in apoptosis.

The Bcl-2 family proteins Bax and Bak are essential for the execution of many apoptotic programs. During apoptosis, Bax translocates to the mitochondria and mediates the permeabilization of the outer membrane, thereby facilitating the release of pro-apoptotic proteins. Yet the mechanistic details of the Bax-induced membrane permeabilization have so far remained elusive.

The Oxidation Status of Mic19 Regulates MICOS Assembly.

The function of mitochondria depends on the proper organization of mitochondrial membranes. The morphology of the inner membrane is regulated by the recently identified mitochondrial contact site and crista organizing system (MICOS) complex. MICOS mutants exhibit alterations in crista formation, leading to mitochondrial dysfunction.

RESOLFT Nanoscopy of Fixed Cells Using a Z-Domain Based Fusion Protein for Labelling.

RESOLFT super-resolution microscopy allows subdiffraction resolution imaging of living cells using low intensities of light. It relies on the light-driven switching of reversible switchable fluorescent proteins (RSFPs). So far, RESOLFT imaging was restricted to living cells, because chemical fixation typically affects the switching characteristics of RSFPs.

MITRAC7 Acts as a COX1-Specific Chaperone and Reveals a Checkpoint during Cytochrome c Oxidase Assembly.

Cytochrome c oxidase, the terminal enzyme of the respiratory chain, is assembled from mitochondria- and nuclear-encoded subunits. The MITRAC complex represents the central assembly intermediate during this process as it receives imported subunits and regulates mitochondrial translation of COX1 mRNA. The molecular processes that promote and regulate the progression of assembly downstream of MITRAC are still unknown.

Mic10 oligomerizes to bend mitochondrial inner membranes at cristae junctions.

The mitochondrial inner membrane is highly folded and displays a complex molecular architecture. Cristae junctions are highly curved tubular openings that separate cristae membrane invaginations from the surrounding boundary membrane. Despite their central role in many vital cellular processes like apoptosis, the details of cristae junction formation remain elusive.

STED super-resolution microscopy reveals an array of MINOS clusters along human mitochondria.

The mitochondrial inner membrane organizing system (MINOS) is a conserved large hetero-oligomeric protein complex in the mitochondrial inner membrane, crucial for the maintenance of cristae morphology. MINOS has been suggested to represent the core of an extended protein network that controls mitochondrial function and structure, and has been linked to several human diseases. The spatial arrangement of MINOS within mitochondria is ill-defined, however.

Rcf1 mediates cytochrome oxidase assembly and respirasome formation, revealing heterogeneity of the enzyme complex.

The terminal enzyme of the mitochondrial respiratory chain, cytochrome oxidase, transfers electrons to molecular oxygen, generating water. Within the inner mitochondrial membrane, cytochrome oxidase assembles into supercomplexes, together with other respiratory chain complexes, forming so-called respirasomes. Little is known about how these higher oligomeric structures are attained.

MINOS1 is a conserved component of mitofilin complexes and required for mitochondrial function and cristae organization.

The inner membrane of mitochondria is especially protein rich and displays a unique morphology characterized by large invaginations, the mitochondrial cristae, and the inner boundary membrane, which is in proximity to the outer membrane. Mitochondrial inner membrane proteins appear to be not evenly distributed in the inner membrane, but instead organize into functionally distinct subcompartments. It is unknown how the organization of the inner membrane is achieved.